Thermostable DNA polymerases are enzymes which have been isolated and recombinantly expressed for a long time since the establishment of the polymerase chain reaction (PCR). However as it is the case for other enzymatical reactions, also the performance of PCR is known to be at least partially hampered by the presence of trace amounts various different reagents such as detergents. On the other hand, the presence of detergents is known to be essential for many polymerase purification protocols and long term stabilization of enzymes in general and DNA polymerases in particular.
Lawyer, F. C. et al. (JBC 264 (1989) 6427-6434) for the first time disclose the cloning and recombinant expression of Taq DNA polymerase. Similarly, U.S. Pat. No. 5,127,155 discloses polymerase formulations which are stabilized with non ionic detergents which are particularly useful for PCR applications. Typical detergents stabilizing detergents used are TRITON X-100 (Union Carbide Chemicals & Plastics Technology Corporation), TWEEN 20 (ICI Americas Inc.) and NONIDET P-40 (Shell International Petroleum Company Limited).
Morever, according to the observations of the inventors of U.S. Pat. No. 5,127,155, the presence of detergents within the disclosed formulations is not only required to maintain enzyme stability, but also to enhance the activity of the polymerase.
Alternative purification methods and formulations have been disclosed. For example, WO 08/077,017 discloses polymerase formulations with anionic and zwitter-ionic detergents instead of non ionic detergents.
Lawyer et al. (PCR Methods and Applications, Cold Spring Harbor, p. 275-287 (1993)) provide improved protocols, wherein the presence of detergents during purification of the enzyme is reduced. Engelke, D. R. et al. (Anal. Biochem. 191 (1990) 396-400) disclose formulations of recombinant Taq polymerase with only trace amounts of detergent, because the finally added storage buffer is free of detergent compounds.
In view of the outlined prior art, it was an object of the present invention to provide an improved polymerase formulation with optimized performance in a polymerase chain reaction (PCR).